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In-plane vibrational modes of cytosine from an ab initio MO calculation
-.
.~ .~
IF&PLANE V5Rii’ICiNAL tiODl?S OF CiYTOSiNE FROM AN AB tilTI MO CALCUIAmON Yoshifumi NISHIMURA FacdryofPitmmacm
_.
i
and Masamichi~TSUBOI
icnlScicncq
(I tzienay
of Tdcp, Hong0 7-3-I.
BInI&o-kl& TOh~-o113.
Japan -_
Received 20 F&mazy 1985
._ Harmonic fora mustants. in-plane vibx=hmal frequaxics.2tnd in-plane 6brational &is of qtosine xs~ d_mlated by-a3 ab initio Hartrcc-Fock SCF MO method_The form mntants were cahhed by the use of an energy gradient method with the 5X0-3G basis set, and then thy were mrrxtai into “4-31G force constants” by the scaling factors given by us previously for the case of uracii The mrrcctcd set of force mnstants can produce a calculated vibrational spectra of cytosine and cytosine1,amino-d~. that can be wdi mrrclated with the observed Raman and~_hfrarcd specua of, these mmpcnmds. wi” little ambiguity_ Thus, the assigamenrs of all *he in-plane viirations arc now practically established The cakuIatcd iiiratioti. modes. in addition. can +cmunt for the mzmtiy published resonance Raman effects of cytosine residue
I_ Introduction For ma&ng vibrational spectroscopy powerful in nucleic acid investigations, it is essential to establish a reliabIe set of normal modes of vibration of each nucleic acid base. For a normal coordinate treatment, in turn, it is essential to ob*%in a reliable set of force constants of. the molecule in question_ In our previous papers [1,2], we showed that it is practical to calculate the force constants of a mclecule, as big as nucleic acid bases, by the use of an ab initio MO method with the STO-3G basis set Then it was p&r&k to predict the vibrational modes of uracil [l] and guanine [2]. by the use. of “4-31G force constants” which. were reached by a correction from the STO-3G force constants_ The calculation could reproduce the observed frequencies, not only of these molecules, but also of their deutemtedtid “N-substituted products_ In addition, it was found that the predicted vibration&.frequ&zies and modes of uracil can well account for its matrix isolation spectrum (IR -and Raman) and that based you our force coristanti~the observed infrared intensities can be well reproduced [3,4]_ -: -..~ (. _ In this paper,, we report a similar study on cytosine By- following the:. established recipe
[1,2,&q we could reach well-characterized normal modes for all ‘of the twenty-three in-plane viirations of this molecule_ This enables Us to make a rather concise set of assignments of the observed Raman Andy infrared bands of cytosine tid its. deuterated product, as will bc presented below_ It will aIso be shown that we can expIain some of the experimental results on the cytosine residue
2. Computational details For the cytosine molecule (fig l), we -used the ~. ab initio SCF MO method with the STO-3G basis set with standard parameters. [7]_ The geometry and the forcccoristants were calculated by using the energy gkdient technique [S] w&h the IMSPAC program [9]_ The optimized geometry isshown in table 1, iu comparison &ith crystallographicahy obtained rksults’[lO]. This should correspond to the geometry’ of the freecytosine molecule, but it shows a g&d coincidence with the averaged vahks : of the bond Iengths and bond angles found. in the X-ray crystallographical analyses of many cytosine derivatives [IO];. except for. a few @ammeters; The gieatest difference (O-084 A) in the calculated and .m experimental bond lengths is fetid for C&+Nx.
_~ ~03~1-0104/85/~03~30 @ &evier Science Publishers B-V. (North-Holland Physics Publishing Division)
.
:
.
: .~
!S-G3G
I’
NJc, c+N,
N,‘G 2: CN, N,H
‘i rz
I.445 1_440
5 ‘k =?
I329 1388 1.019 cz20 1391 1.012
5 G
=,o :zo
=s ‘io r,
Z’,
‘11
1.309 IA71
discrepancies in the bond an&s are rather great For N&C, and C&N, (43” and -5-O’=. respeetiveIy as shown in table I)_ It is tmderstandable that the local geometry around the N&N, part of the mokeule is most susceptiiIe of intermokcttIar interactions in crystals_ Such gcometriCal differences are actttahy found among different crystals of eytosine derivatives [lo]. Due allowance should therefore be given later in comparing the observed spectrnm of crystahine cytosme with that eaktdated on the basis of this optimized geometryForce constants of the in-plane vibrations were caIcnIated by numerical differentiation of the gradients (Forces) in the eartesian coordinate system and then transformed into an internal coordinate system_ The internaI coordinates S adopted here are shown in table 2_ The redundancy of the angtdar coordinates caused by the ring system has already been removed_ These STO-3G Force co= stants of eytosine were then eorreeted into “4-31G force constants”_ The ?XO-3G to 4-31G eonversion factors were obtained previously [l] by a &met eompaxison of the actuahy eakukted force constants both with the SC?-3G and 4-31G basis sets for mea., acrylaIdehyde and Formamt‘de In transferring these conversion factors into cytosin~ onr rest&s of force constant eaIaJIations of imidazole wi*& the STO-3G and 3-2lG basis sets were ako taken into consideration The eon-version The
r13 2:
122-8 1.087 116.7 1172 126-I 117.0 119-9 117-l 1195 1233.6 1169 116.8 1203 1226 123A 116-4 119-9 121.4 119.5 119.1
z:: BIS
=A&
HN,C, oc2Ni G&N,
z!I 2
WC&'9
is3
CSCJ% HC;C, CC,H HC.SC, N&H HNIC, ‘=PsJ-%o HnN& H,oN,H,,
2
bJ
_I399
1x6 1I.334 I.426 1337 1364 1237 1337
1.013 I.076
liz z&. N&N; CzN,C, WAC, C&c,
Ok
2 2 2 az
=’ For numbering of dis.kBindv bJ F&f_ [lOJ_
120-6 118-9 120_0 121.8 117-6 121.0 1192 I219 117.9 121s
anda&esseefig_l_Unitszrin
Factors cii nsed For the diagonal elements ci of eytosine are given in the last column of table 2, where Ei(4-31G)
= c~~~~(STO-~G).
For each oFFdiagonaI element $, Factor cii Was assumedto6egivenby
(0 the eon-e&on
Here, howe& very sn.talI off-d&ona.I &zmen& were ignored, because they are not effective at all Thus, the stretehing+retching elements smaller
.:r
i
si ”
c,
N,G_ str-
s, = Jr,
0.96
C-N3
SW
%=4r,
096
N,C,
str.
Gc, QrCN, str-
-0.96 o_82
&==b6 S,=4r, &=4r, &=4r,
N,H str_ c,o SQ-_ c,Ng str. N,H,, SW N,Htt SWGH str_ SH str_ ring dcf_ I ring dcf_ II ring dcf. III N,H Lund_ c-0 bald C,N, bend qtIbald_ C&H bend_ NH2 sciss NH2 rock
... . ,
_
o-w_
Jj=ar, ;s, = 4r. %=4r..
Gc, SW
._
GO
=
450
St, St, S, s,, s,,
= = = = =
Art, 4riz 4r,, 4&, - 4B,5 •!-4&s -
&6
= 4&,
-
24i31, -
4bq5
-
4i317 +
4i316 +248,?
4BM + 4&a -
4&s -
4Bi9
-.4h
-
4i3,,
4B19
S*,-4&m-4&9 S,, = 4&_, - 413, St, = a/323- 4&z s&=4&,-4s.55 &,=4&‘4& L_ = 24S, - 4ht - 4ao s; = a#.&0- 4&,
O_% 0.87 OX? 0.96 0.87 0.87 0.82 OS2 1.08 1.08 1.08 O-91 1.08 1.06 091 0.91 0.91 1.0
_---
;.~ i .~.
.m.
:
a’ For numbering of distmas and angks see fig. 1.
0.1 mdyne/& the stretching-bending elements smaller than 0.03 mdyne, and the dements _smaller than 0.01 bending_-_bending mdyne A were all assumed to be zero. ‘The final set of F matrix elements arc given in table 3. The viirational frequencies and normal modes of v&rations were calculated from the force constants (table 3) by a program written by us, which is a modified version of NCTB program written by Shimanouchi Ill]_ The normal modes have been shown here by a plotter output of the calculated Lx matrix, than
x =.L$,
_-
_
a-
Where~X 1s *e cartestan dnp*acem~ent &5ordinaZ (Ax and by for each of the thirteen atoms) vector and Q is the normal coordinates (twenty-three) vector_ Note that elements of Lx have a dimension PL.Lx was writof m&s-r/l_ The piotter prod ten by Dr_ Y_ Sugawam, and kind& provided by
her_
I:
:~--
__
: .--
: ~-
3_ Results The cakulated normal frequencies of cytosine and cytosine-1, amino-d3 are given in table 4 and are graphically presented in fig 5 along with the Baman and infrared frequencies reported by Susi et al [X2] for the crysWhne powders. The c&ulatcd normal modes are shown in fig 3 for cytosine and in fig 4 for.cytosine-1, amino-c&_ The vibrational modes are ako indicated by symbols (v,“, Tr, -etc.). in table 4 and in fig& 214:. These symbols represents %ymmetry coordinates’.’ ~of a_ hypotheticaBy symmetrized Lqtosine molecule; wh_ose six-membered ring, for example, is a regular hexagon [!3]_LSomc of -such standardized viirational modes are shown in. fig_ -5_ Every actual .’ vibmtional- mode: is a-~blend of a- few of tbesc : idealized modes, but oftenthe’contrihution~of one : -of- them looks predo-mmant in it :.It wouId be convenient to. designate each a&& mode. under thesamenameofsu~apredominantmode--:..~--:. Because_ we are dea@g: only with: the-in-plane. .-
:
:
2 3 4
5 6 7 8 9 ICI 11
12 13 I4 15 16 17 IS 19 20 21 Ic 23
0554
--a357 O_lOd -0-361 0597 o-0 I_014 o-0 01) 0.0 o-0 0.0 O-159 0119 o-0 o-0 0534 0.0 0_035 O-042 o-0 0.0
627.5 033 -0310 0313 -0260 a0 0.853 -022 o-0 0.0 o-0 0.0 0328 -0332 -0-412 0.0 -0-434 0.0 -0-032 a0 01) 0.0
o-669 -0_311 0.4x a0 -0.179 0.994 0.0 0.0 0.0 o-0 0.411 -0206 -0570 -0_043 O-057 OJXS o-0 O-0 o-0 O-071
6532 O-697 -0-614 0.0 01) 0391 0.0 0.0 0.0 01) QICt4 03 02&l O-053 -0_056 -0-327 0.179 0.0 o-0 o-0
vibrations of cytosine in this study, only the Raman and infrared bands assignable to the in-plane vibrations are given in table 4 and in fig 2 We can distin,puishbetween in-plane and out-of-pIane vibrations based on our present caktdation Our distinction is mostIy the same as that propo&d by Susi et al (12_14&Thus, markedIyb-mad and strong infrared bands at 823 and 520 cm-’ of cytosine and those at 605 and 388 cm-’ of cytosinel.amino+ are reasonably assigned respectively to NtH out-of-piane bending, NH2 wagging NrD out-of-plane bending, and ND_ wagging viirations With referenceto an anisotropy study of the infrared absorptions in a single crystaI of l-methyltbymine [15].568 and 442 cm-’ bands of qtosine and 550 and 438 cm-t bands of cytosinel,aminti3 are -ascribed to out-of-plane modes_ These are alI removed in table 4 and in fig 2 In additioa,twowetakRamanbands atlOlOand994 _-’ of cytosine and two Raman bands at 1046 and 1005 cm-t of cytosine-l,amincAs conId be assigned to CH out-of-plane viirations_ Therefore, th~WeredSOremove&
O-743 -0.097 -0.126 o-0 0.0 0.0 01) 01) 0.0 -0342 0.0 O-0 0_055 O-044 -O_lS-J O_I20 0.0 0.0
s_4i3 o-0 o-0 -0-149 o-0 0.0 --a103 o-0 -0_192 0.106 -0337 -O_l% -0.137 -0.065 0.0 -0113 0.0 0.0
5.169 0.0 0.0 O-0 o-0 o-0 o-0 o-0 0.0 o-0 o-0 O-0 0.0 0.0 0.0 o-0 0.0
13347 o-0 0.0 0.0 o-0 0.0 0175 0.199 -0308 0.0 0.077 0.0 0.0 0.0 o-0 0.0
8.452 o-0 o-0 0.0 0.0 0.186 -0302 0.0 o-0 -0_06s 0.072 0.0 o-0 -0_170 0.061
S-453 -a192 0.0 0.0 0.0 0.0 0.0 01) o-0 0.079 0.0 o-0 0.055 O_OS6
By an extensive test of the ab initio force constants [l-3,5-6,8.16-21). it is now generahy claimed that the 4-31G force constants of a moIecule can predict a correct in-plane vibrational mode spectrum_ When the calculated modes of a molecule are arranged in the decmasing order of the calcuhxted frequencies, along the arrangement of its observed fundamental bands in the decreasing order of the observed frequencies then one can reliably comxct one-by-one the members of these two arrangementswithout any cross connection- It is true that each of the caLdated normal frequencies comes out to be lo-20% higher than the corresponding observed fundamental frequency. Thisis partlybecauseoft!~eanhaxmonicityofthe vibration and partly &cause of the inability in the present MO calculation to incIude proper configuration interaction [20]. Nevertheless. the viirationaI modes. isotope effects, and infrared intensities of a moIecuIe are considered to be predicted by the 4-31G HF SCF MO method with a significant pre&iotL Based upon such a -generaI reliability of the
8.435 0.0 0.0 0.0 o-0 o-0 O-0 0.0 -0.059 0.0 o-0 0.03s -0.072
6.337 0.0 O-0 o-0 o-0 0.0 o-0 0.0 o-0 o-0 o-0 0.0
6.173 0.0 0.0 0.0 O-0 o-0 0.0 0.0 0.0 o-0 0.0
l-843 o-0 o-0 0.0 0.035 0.0 0.0 0.0 o-0 o-0
1.729 o-0 o-0
0.173 -0.045 0_087 -0_096 o-0 0.0
1.740 0.132 0.0 -0.119 0.059 o-0 o-0 0.0
o&O1 -0.060 0.0 O-014 o-0 o-0 0.0
1336 -0.041 0.0 -0.027 o-0 0.0
1.352 0.0 -0.020 0.036 0.193
0590 0.0 0.0 0.0
0.635 a.0 0.0
0574 -0.014
0.693
4-31G
calculation, the observed Raman and &Ifrared bands are assigned as shown in fig_ 2_ There
stretching vibrations, respectively_ In the 1800-850 cm-’ regiow the highest two
are a number of aiterations necessaq from the assignments proposed by Susi et al. 1121. Two strong infrared bands of cytosine observed at 3380 tid 3169 cm-’ are assignable to the NHI antisymmetric and symmetric stretching vibrations, respectively, and those of cytosine-dz at 2545 and 2337 cm-’ to the ND2 antisymmetric
observed frequencies, 1694 and 1653 cm-‘, are to be assigned respectively to the Cz=O and G% (or v”,) stretching vibrations_ Our cakulation indicates that the next highest frequency should be that of the NH, scissoring motion, and the strong infrared band at 1615 cm-’ corresponds to it_ It is
and symmetric stretching vibrations, respectively_ A fairly strong Raman line at 3117 cm-’ and a
moved down to 1182 cm-’ on deuteration- The next three highest frequencies should be assigned to v,” of the hexagon, C,-(NH,)
stretching, and
changed on NI and amino deuteration and are assigned to two CH stretching vibrations. Our calculation indicates that the N,-H stretching vibration should be Ioca~&_ at + slightly higher frequency than that of the NH, Metric stretch-
N,H in-plane bending vibrations in turn; and these are correlated respectivelywith the obserred 1533, 1498, and 1462 cm-’ Raman lines. On deuteration, the fit two remain nearly unchan~~ &iIe the last one (N,H b&d_) is moved to 950 ~cm-.‘_ The cakulation instructs-&at the GH.- GH in;
ing vibration in an isolated molecule_ It k.know& [22]. however, that a hydrogen-bonded irpino group gives often a broad kand at a much lower frequency. The broad infrared band of ‘cytosine around 2700 cm-’ and that of cytosine-d, around
ph&e bending vibration should come next, followed by the KekulC vibration, and next the GH GH lSO”-out-of-phase bending Correspond+g to these, w prominent Ramv Iinks are observed &t 1361, 1276, &id 1247 cm-‘, req+ctiv~ly._ T+
weaker one around
2150 cm-’ -
3050 cm-’
both remain un-
can be assigned ;o the N,H and wID NH, -. ~.I. ~. . .:-_ ~.~. .: .I
ro&ng vibration is predic+i to be the r&t _:_
Coicd.
Il. id 1
L
-_
-a
l-__ -_
l__
__
__
-_
s_ -_
y
.E
Obsi
E
c
R,R
--_
--___ --__ --_-_
_s-
__-
-
-_
:
S.
.
--.
. :
;-,
II
.I
-
--
:=__
:
:
II
a
I
I’i ‘I .
I
I
.
s . . . --. .-_
:-t__
-_z_=_ :
-_
Ill
I .
I
I L
.
:
iI
I
I II
I
9
T
2z
2
1864
1782
Cl633)
Cl6l2)
1645
Cl4981
I575
(14621
1083
(971)
_
639
(5971
.. . .
381
t4001
._
--.
1699
344'(369)
(1514)
highest
one, and the ll&
.&I-~
Rkxan Iin
assignabIetoit_.The threelowest-frequencyyibqtticnsin the 1800-850 cm-' range ai 1108,_971, T@
@
WI2 tot0
and 894 cm-’ are assignable to three-r&g vii& : $ons, v&_ Tr, ana v&, i&pec+Iy_ 1-nr+se three ,~ vibrations, pronounced couphngs unth the -NH, r&g mqtion are found (see -thecalculated 1197,
;:,1485
--
-
ring vibrations, On deuteratiqn, the an+io rocking vibration comes d&k to 850 cm-’ (fairly-s,trong infrared band),- and. a& is shifted up to 1021. a+-*_ In ad&ioq. v& is pushed up. 3~ hi&; as .. 1383 _cm-:_ throkgh +_mu@ing with the N,D hepd; ., .~ing niotioi -Itie ~K@ctxl* motion, 0; ,+:~othei_~ up 06 deute,mti&, fro@. ..
‘.
.-
...
-, :.
:
._.. . . .-
_ ~.._ ...._: _.. --
3 4 5 6 7 8 9
4013 3sM 3803 341s 3375 193il IS64 US2 1722
IO 11 12 13 14 15 16 17 18 19 20 21 22 23
Ia5 I575 1476 1364 1309 1241 1197 19S-s 988 S2L 639 572 567 3SL
1 2
NH,
PSU-
3354
N,H* NH, s str GHSW GH= c=o sv q=c,str NH2 sci d &N Str 8N,H 6CH+ K&d& 6CHNH2 r G”, -r&-i da bmtb $ sbm SCN
_ _
3176 3117 3050 1694 3653 1612 1533 14% 1462 1361 1276 1247 1x48 1108 971 894 792 597 546 533 400
3350 2700 3169
1703 1662 16i5 1538 1505 1465 1364 it77 1236 1150 1100 966 793 600 549 533
3418 3375 2978 2833 2751 1923
2388 1648 15% 1514 1491 1383 1316 1282 11% 1130 1021 965 --
1855 1699 1653 1521 1444 1366 12SO 1259 1123 1@aS1aM IsG9 Sll
2545 2150 2337 1637 1605 1517 1505 1377 1316 12Sl 1182 1120
849 m 590 524 504 369
628 555 541 344
=’ E$Ihcuscofrtief--~tsgi~znintile3_ ” Rcf_ [9f
tiv&y_ All of these Four are moved narrowly towards lower frequency on N, and amino deuteration_
4, IxscusGon In our present study- the C,-amino stretching Frequency (1505 cm-‘) of cytosine is Found to be of interest_ This is extraordimniIy high in comparison, For exampI< with the C-amino stretchingfrequency (1276 cm-‘) of anihne [23]. This C-N bond is considered to have the greatest doublebond character among the C-(NH,) bonds of nucleic acid bases (cytosine, adenine, and guanine) [24]_ The height OF the barrier in the >C-NH, internal rotation decmases in going from cytosine, to adenine, and guanineAnother point OF interest in the cytosine mole-
cule
is its
“Kekdi
structure?_
As is seerk in the last
of table 1, the six bonds Forming the ring are long and short alternately. The bond-stretching Force constant changes also up and down ahernately on going along the six-membered ring (see the first six diagonal elements shown in tabIe 3) Thus, the bond order is considered to become higher and lower on going along the ring ThereFore, the KekuIi viiration (Kk), in which every bond in the six-membered ring stretches and contracts with a 180° phase difference from the stretching and contraction of the adjacent bond, may cause a gre& over&I poIari%abiIity oscihation in cytosine, whereas it should never in benzene_ Actually, the Raman line at 1276 cm-’ assigned to Kk of cytosine is very strong (fig 3) with the 488.0 nm excitation. The assignment of Kk in the cytidine residue is not made as straightForwardly as for cytosine itcohmn
.
-
:
-.
self_ -In-the Raman -.spectrum~of 5’CM.p
or 5’-
dCMP, there are two Iin&-at 1290 and.1250 cm-‘, and.& a. broad band %&nd 1230 Cm-‘. The band a&md 1230 cm-1 is .as&nabie -to 6CH-. The fint two Raman lines are considered to be originated by the &In-ationaI coupling between the Kk .motion. of the cytosine moiety -and a sugar vibration. Considering~ the intensities of the&e two Raman Ii&s in the visible excitation, however, we prefer to assign the 1250 cm-’ Line to a mode in which Kekule motion is predominant. The KekuIe structure of cytosine in its ground electronic state must .be-disorganiz& -in its first electronic excited state at 268 nm [25]_ As is seen in fig_ 6, on going from the highest occupied to the lowest vacant molecular orbit& the Ns=C, and Cs=C, bonds should change from bonding. to antibonding; while the C,-N, and C,-C, bonds should change from antibonding to bondinK, the bond order decreases in the former two and increases in the latter two, and therefore G-N, must be shortened, N,=C~ lengthened, C,-C, shortened, and C,=C, lengthened_ Thus, the geometry change in the lo&st electronic transition may rather match the Kekule motion_ Therefore, the Kekult vibration around 1250 cm-’ of the cytosine residue must be active in its resonance Raman spectrum with excitation in its 268 nm absorption band. This is found to be actuahy the case. The 1243 cm-’ Iine of S-&BP and the 1255 cm-’ Iine of S-dCMP are very strong in their Raman spectra excited at 257.3 run [26,27] or at 266 nm [28-301. It is noteworthy, on the other. hand, that they are completely absent with excitation at 240, 218, or 200 nm [28-301. The. geometry of the cytosine residue in the second- and the third-lowest excited states does not. seem. to match the KekuIi type deformation, The NH, and ND, scissoring viirations have been Iocated in our present study at -1612 (cytosine) arid 1190 (cytosine-cZs) cm-‘, respectiveIy_ This supports the assignment of the 1607: cm-’ Raman Iine of 5’dCMP (in aqueoussolution) to its NH, scissoring .vibmtion made recentIy by Fodor et al [30]. To its ND, scissoring, we assign a Raman Iine at 1181 -cm-?-of S-dCMI? in l&O sohrtion~ The 1607 ‘-cm-l line. ‘of 5IdCMP w& found to be enhanced by ihe 200 inn excitation
transition from a u orbital to the 3s orbit$;of&e' nitrogen atom around 2O(I run. Such a’transitiori_.. should have its transition moment _&mg the-dir@. tion (2) perpendicular to the rnol&rl&~pIa&<~~@id me the zz component of the Raman sc%tteririgte&& 1 of the NH, .scissoring vibration mi&t-be m-r&~:-
n&e -5th &is &&o~c t.ra,n.sition_.-. _-Z-~-.ci ._:- _I’ -As discuss~ above, the present viirationd assignmentsfor the cytosine molecule &e often ;Seful in the vibrational &gnments for & cytidine derivative_ S-dCMP, for example, shows -weaker Raman scattering in the low-frequency &de of the strong 1255 cm-’ Iine (Kk) ~withpeaks at 1242 ’ and 1214 cm-’ 1301.The 1242 cm-’ is assignable to SCH-, but the 1214 cm-’ line‘& not attributabie to any_ of the-vibrations of the-base moiety. ] Hence this is suggested to be caused by. the ‘de_ oxyribose moiety_ As mentioned in connection with the assignment of the Kekule vibration above, we may expect a few vibrations of the sugar in the 1200-1300 cm-’ range_ Indeed, we have recently found the spectraI feature in the 1300-1200 cm-’ range of cytidine is sensitive to the. deoxyribose conformation_ Some vibrational couplings between them would cause such a conformation dependent
Fig 6. ‘Ihe highest occupied and _lowes< va&ant ~m&klar orbita& of the P el&ns in thecytosine mokcul~ on the basis of & ~publi+hed part of the result&of a P&iser+axT-Pople calculation made by Nagata et al [25]_ Open w$ closed .tie~ idicace that the upper bra&he+ of the ato’&& orbit+ of the.= ekctrons iic &sitive And nk@iv~$n+ectively_ l-li~_s+e of the &de indicates thk appr&+ate value of the.coetXcia;t of th+ par&lar atomic orbital in .&e relevant moIe&+~ital. a&zre (rad;u~)~ E coefkitnt .~
~_ ~.:
-.
spew-al feature- The details w-ill be given in our comingpaper-
Our. sinaxe thanks are due to Professor K. Morokuma, Institute for MoIecuIar Science, and Dr_ S_ Kate, Nagoya Utiersity, for their kind arrangements and instr~.~ctio~~~ in our ab initio MO cahhion with the ILMSPAC progg Tf?e normaI frequency and normal mode caiadation was carried out at the Computer Center of IMS (HIIAC M-ZGOH)_ The pIotter output of the normal modes was made at the Computer Center, University of Tokyo (HITAC M-20OH)_ This work u-as supported by a grant (No_ 57060004) from the ~finistry of Education. !Zciace. and Culture of
Japan References Y_ Pikhimm
M -E-sub& S Karo and K Mot-&w J_ Am Cbem_ Sac 103 (1951) 1354. Y_ Nisbimum_ M_ Tsuboi S Kate and K_ Morokuma_ B&L Cbun k Japan 58 (1985) 638_ Szcwni& AS-L Now& K Rostkounlia. K_ SnuL W-B_ Pesso~ and D. Sbqar. J. Am. Cbcm Sot 105 (1953) 5969; SChkLScotrK_Szcqx&&andW_B.Pason.J.Am_ cban sac 106 (19&I) 3415_ _A_LBymsMAStucke);aIkdl_ICG&specvochirn Acct 40.~ (1934) 41s hLTsuboizndY_Nichimunin:Ramanspcruoscop_vno&mar. rdr J. kicombc and P-V_ Huoq iinurmld (Wdey_ New York 1952) p_ 683_ Y_ Nisbimun ;K Tsuboi. S. Kate and K rhkrokuma. in: y-linearandmmlin~_&J_ R==Qspecvavop ksannbe and P-V_ Huoq (Wiley. New York 1982) p_ 703_ SJ. Hcbrc. RE Stewart and J-4 Pople J_ than Phyx 51 (1969) 2657_ P_ Pulay. in: AppGceJn of ekuonic z3mnxmc dumy. cd_ H_F_ Scbacfcr III (Pknum Rcss. New York. X977) ch. 4. K Sf&p s Kate_ Ecxifaura. I_ OtlmiltG s_ skai a.ad S OLxm. IhfS Computer Caxtcr Liiraq program. x0_ 0372 @9scQ_ R T&or and O_ Kenn;rrb J. hfoL Srruh 78 (l9SZ) 1_
for nd coordiT_Sbksw&_Computerp~ natc weatnwnL of poI~icomic mokadcs rqanmalt OF Cba&ary_ plnkn-si~ of Tokyo. Tokyo. Jqxme KSu&JSAniandJMPmc&.SpawcbkAcmWA (x973) 72s
EB. Wu J-C Decks &nd PC Cross, Mokcuhr C%ricions (McGxau--Hi& -New York. 1955) ch. 10. [X4] I-L Susi &xi JS Ard. spactrochim_Aaa 27A (l97l) 1549_ [15] Y_ Krogohs S HigudC and U Tsuboi. Spearoctrim Acta 23A- (l%7) %9_ [la] Y_ Sugawaxa. Y_ Hamada. AY_ ?&&a. ti Tsboi, S Kao and K_ Morokxka. QmPhk 50 (l980) 105; .Y_ Suga=w AY_ Hirakamx, M_ Tsuboi S Kate and i MB C&an mk 62 0981 j 339; Y_ w N_ Tanaka. Y_ Sugawam AY. Hi&k& I$_ Tsuboi S Kate and K Meti J_ MoL Spectry_ 96 (1982) 313_ 1171 CL Blom_ C Akona and Ckkam, MoL Phys 34 &I?) [l3]
557; CJZ BIom and C Alcona. CE
hid
Phys 34 (I-
177:
Bkm. i_P_ Otto and C _AImna.MoL Phyr 32 0976) 1137; CL Bkxn and C Altozu MoL Pb_yc 31(1976) lm; CE Blom PJ_ .S&q&md and C Alto= MoL Pb= 31 (1976) 1359. [lSl C_W. Bock. PmGeorge and hi_%ad~tmiu~J. MoL Spear-y_
7s (1979) 248;
CW_ Bock, M. Tmcband P_ Gcorgc J. MoL Spatry_ SO (1980> 131; 84 (19sO) 243.2s [191 f_ PkIay and W_ Meyer. J_ hid Specu-y_ 40 (197l) 59; W_ Meyer and P_ P&y. Theoret Chin Acta 32 (1974) 353: P. Ptday and F. -I’&&. J. Mol. Struct 29 (1975) 239; P_ P&y_ G_ Fogamsi. F_ Pang and J-E- Boggs J_ Am_ Cban Sot 101 (1979) 2550: P_ Pulay. G_ Fogam& G. Pongcr. JE Boggr and h Vzqa, J_ Am Cbcme Sot 105 (1983) 7037_ [201 N_ Tanz&a. Y_ I-Izmzda. Y_ Suga~swa. kM_Tsuboi S Kato and K_ hfot-okxmsa. J. MoL Spatry_ 99 0983) 245; N_Tanaka,Y_H-daandM_Tsuboi,Ckm_Pbyr94 (1985) 65. [IIf Y_ Sugaaarq A_Y_ Hirakawa and M_ Tsuboi, J_ MoL spa=uy_ 10s (1984) x6_ 1221 CL An@. I Chau Sot (l%l) 5w. 1231 M_ Tsuboi. Spafmchk Acta 16 0960) 505_ [24] M_ Nakau&i. S. Tstmcda. S. TakahasE and hi. T_suboi.J. Biodrem (Tokyo) 97 (3985). to be published_ (251 C Nagam. A lmamura and H Fujira in: A&anccs in biophysics. VoL 4. aL M Kotani (Uniuazity of Tokyo Press Tokyo. ‘-mj pm1. [26] Y_ Nkbimma. AYe Hirakana and U Tsuboi. C&em_ LcIKux(l977.)907_ [2q Y_ Nishimm-& A_Y_ m and M Tsuboi, in: AdtanccsiItinfrarrdand~~roscopy*voL5,cds_ RJ.Ha Clark and RE Hcncr (Heyday, London, 1978) p_ 212 1281I-D. Zicglcr. -B Hudson. D-P. Screw and W_I._ Pc?icok!s,Biopolymas 23 (1984) 2067_ [zsl WX Kubasdc. B_ Hudson and WE PcticoIas, Pmc NatL ~Acxd.Sci_US.cobekpublisbed. [30] S_P+ Fodor, RP. Rava aiid T-G: $& J_ AI+ C+_ sot 107 (1985) 1520_
.~ .~
IF&PLANE V5Rii’ICiNAL tiODl?S OF CiYTOSiNE FROM AN AB tilTI MO CALCUIAmON Yoshifumi NISHIMURA FacdryofPitmmacm
_.
i
and Masamichi~TSUBOI
icnlScicncq
(I tzienay
of Tdcp, Hong0 7-3-I.
BInI&o-kl& TOh~-o113.
Japan -_
Received 20 F&mazy 1985
._ Harmonic fora mustants. in-plane vibx=hmal frequaxics.2tnd in-plane 6brational &is of qtosine xs~ d_mlated by-a3 ab initio Hartrcc-Fock SCF MO method_The form mntants were cahhed by the use of an energy gradient method with the 5X0-3G basis set, and then thy were mrrxtai into “4-31G force constants” by the scaling factors given by us previously for the case of uracii The mrrcctcd set of force mnstants can produce a calculated vibrational spectra of cytosine and cytosine1,amino-d~. that can be wdi mrrclated with the observed Raman and~_hfrarcd specua of, these mmpcnmds. wi” little ambiguity_ Thus, the assigamenrs of all *he in-plane viirations arc now practically established The cakuIatcd iiiratioti. modes. in addition. can +cmunt for the mzmtiy published resonance Raman effects of cytosine residue
I_ Introduction For ma&ng vibrational spectroscopy powerful in nucleic acid investigations, it is essential to establish a reliabIe set of normal modes of vibration of each nucleic acid base. For a normal coordinate treatment, in turn, it is essential to ob*%in a reliable set of force constants of. the molecule in question_ In our previous papers [1,2], we showed that it is practical to calculate the force constants of a mclecule, as big as nucleic acid bases, by the use of an ab initio MO method with the STO-3G basis set Then it was p&r&k to predict the vibrational modes of uracil [l] and guanine [2]. by the use. of “4-31G force constants” which. were reached by a correction from the STO-3G force constants_ The calculation could reproduce the observed frequencies, not only of these molecules, but also of their deutemtedtid “N-substituted products_ In addition, it was found that the predicted vibration&.frequ&zies and modes of uracil can well account for its matrix isolation spectrum (IR -and Raman) and that based you our force coristanti~the observed infrared intensities can be well reproduced [3,4]_ -: -..~ (. _ In this paper,, we report a similar study on cytosine By- following the:. established recipe
[1,2,&q we could reach well-characterized normal modes for all ‘of the twenty-three in-plane viirations of this molecule_ This enables Us to make a rather concise set of assignments of the observed Raman Andy infrared bands of cytosine tid its. deuterated product, as will bc presented below_ It will aIso be shown that we can expIain some of the experimental results on the cytosine residue
2. Computational details For the cytosine molecule (fig l), we -used the ~. ab initio SCF MO method with the STO-3G basis set with standard parameters. [7]_ The geometry and the forcccoristants were calculated by using the energy gkdient technique [S] w&h the IMSPAC program [9]_ The optimized geometry isshown in table 1, iu comparison &ith crystallographicahy obtained rksults’[lO]. This should correspond to the geometry’ of the freecytosine molecule, but it shows a g&d coincidence with the averaged vahks : of the bond Iengths and bond angles found. in the X-ray crystallographical analyses of many cytosine derivatives [IO];. except for. a few @ammeters; The gieatest difference (O-084 A) in the calculated and .m experimental bond lengths is fetid for C&+Nx.
_~ ~03~1-0104/85/~03~30 @ &evier Science Publishers B-V. (North-Holland Physics Publishing Division)
.
:
.
: .~
!S-G3G
I’
NJc, c+N,
N,‘G 2: CN, N,H
‘i rz
I.445 1_440
5 ‘k =?
I329 1388 1.019 cz20 1391 1.012
5 G
=,o :zo
=s ‘io r,
Z’,
‘11
1.309 IA71
discrepancies in the bond an&s are rather great For N&C, and C&N, (43” and -5-O’=. respeetiveIy as shown in table I)_ It is tmderstandable that the local geometry around the N&N, part of the mokeule is most susceptiiIe of intermokcttIar interactions in crystals_ Such gcometriCal differences are actttahy found among different crystals of eytosine derivatives [lo]. Due allowance should therefore be given later in comparing the observed spectrnm of crystahine cytosme with that eaktdated on the basis of this optimized geometryForce constants of the in-plane vibrations were caIcnIated by numerical differentiation of the gradients (Forces) in the eartesian coordinate system and then transformed into an internal coordinate system_ The internaI coordinates S adopted here are shown in table 2_ The redundancy of the angtdar coordinates caused by the ring system has already been removed_ These STO-3G Force co= stants of eytosine were then eorreeted into “4-31G force constants”_ The ?XO-3G to 4-31G eonversion factors were obtained previously [l] by a &met eompaxison of the actuahy eakukted force constants both with the SC?-3G and 4-31G basis sets for mea., acrylaIdehyde and Formamt‘de In transferring these conversion factors into cytosin~ onr rest&s of force constant eaIaJIations of imidazole wi*& the STO-3G and 3-2lG basis sets were ako taken into consideration The eon-version The
r13 2:
122-8 1.087 116.7 1172 126-I 117.0 119-9 117-l 1195 1233.6 1169 116.8 1203 1226 123A 116-4 119-9 121.4 119.5 119.1
z:: BIS
=A&
HN,C, oc2Ni G&N,
z!I 2
WC&'9
is3
CSCJ% HC;C, CC,H HC.SC, N&H HNIC, ‘=PsJ-%o HnN& H,oN,H,,
2
bJ
_I399
1x6 1I.334 I.426 1337 1364 1237 1337
1.013 I.076
liz z&. N&N; CzN,C, WAC, C&c,
Ok
2 2 2 az
=’ For numbering of dis.kBindv bJ F&f_ [lOJ_
120-6 118-9 120_0 121.8 117-6 121.0 1192 I219 117.9 121s
anda&esseefig_l_Unitszrin
Factors cii nsed For the diagonal elements ci of eytosine are given in the last column of table 2, where Ei(4-31G)
= c~~~~(STO-~G).
For each oFFdiagonaI element $, Factor cii Was assumedto6egivenby
(0 the eon-e&on
Here, howe& very sn.talI off-d&ona.I &zmen& were ignored, because they are not effective at all Thus, the stretehing+retching elements smaller
.:r
i
si ”
c,
N,G_ str-
s, = Jr,
0.96
C-N3
SW
%=4r,
096
N,C,
str.
Gc, QrCN, str-
-0.96 o_82
&==b6 S,=4r, &=4r, &=4r,
N,H str_ c,o SQ-_ c,Ng str. N,H,, SW N,Htt SWGH str_ SH str_ ring dcf_ I ring dcf_ II ring dcf. III N,H Lund_ c-0 bald C,N, bend qtIbald_ C&H bend_ NH2 sciss NH2 rock
... . ,
_
o-w_
Jj=ar, ;s, = 4r. %=4r..
Gc, SW
._
GO
=
450
St, St, S, s,, s,,
= = = = =
Art, 4riz 4r,, 4&, - 4B,5 •!-4&s -
&6
= 4&,
-
24i31, -
4bq5
-
4i317 +
4i316 +248,?
4BM + 4&a -
4&s -
4Bi9
-.4h
-
4i3,,
4B19
S*,-4&m-4&9 S,, = 4&_, - 413, St, = a/323- 4&z s&=4&,-4s.55 &,=4&‘4& L_ = 24S, - 4ht - 4ao s; = a#.&0- 4&,
O_% 0.87 OX? 0.96 0.87 0.87 0.82 OS2 1.08 1.08 1.08 O-91 1.08 1.06 091 0.91 0.91 1.0
_---
;.~ i .~.
.m.
:
a’ For numbering of distmas and angks see fig. 1.
0.1 mdyne/& the stretching-bending elements smaller than 0.03 mdyne, and the dements _smaller than 0.01 bending_-_bending mdyne A were all assumed to be zero. ‘The final set of F matrix elements arc given in table 3. The viirational frequencies and normal modes of v&rations were calculated from the force constants (table 3) by a program written by us, which is a modified version of NCTB program written by Shimanouchi Ill]_ The normal modes have been shown here by a plotter output of the calculated Lx matrix, than
x =.L$,
_-
_
a-
Where~X 1s *e cartestan dnp*acem~ent &5ordinaZ (Ax and by for each of the thirteen atoms) vector and Q is the normal coordinates (twenty-three) vector_ Note that elements of Lx have a dimension PL.Lx was writof m&s-r/l_ The piotter prod ten by Dr_ Y_ Sugawam, and kind& provided by
her_
I:
:~--
__
: .--
: ~-
3_ Results The cakulated normal frequencies of cytosine and cytosine-1, amino-d3 are given in table 4 and are graphically presented in fig 5 along with the Baman and infrared frequencies reported by Susi et al [X2] for the crysWhne powders. The c&ulatcd normal modes are shown in fig 3 for cytosine and in fig 4 for.cytosine-1, amino-c&_ The vibrational modes are ako indicated by symbols (v,“, Tr, -etc.). in table 4 and in fig& 214:. These symbols represents %ymmetry coordinates’.’ ~of a_ hypotheticaBy symmetrized Lqtosine molecule; wh_ose six-membered ring, for example, is a regular hexagon [!3]_LSomc of -such standardized viirational modes are shown in. fig_ -5_ Every actual .’ vibmtional- mode: is a-~blend of a- few of tbesc : idealized modes, but oftenthe’contrihution~of one : -of- them looks predo-mmant in it :.It wouId be convenient to. designate each a&& mode. under thesamenameofsu~apredominantmode--:..~--:. Because_ we are dea@g: only with: the-in-plane. .-
:
:
2 3 4
5 6 7 8 9 ICI 11
12 13 I4 15 16 17 IS 19 20 21 Ic 23
0554
--a357 O_lOd -0-361 0597 o-0 I_014 o-0 01) 0.0 o-0 0.0 O-159 0119 o-0 o-0 0534 0.0 0_035 O-042 o-0 0.0
627.5 033 -0310 0313 -0260 a0 0.853 -022 o-0 0.0 o-0 0.0 0328 -0332 -0-412 0.0 -0-434 0.0 -0-032 a0 01) 0.0
o-669 -0_311 0.4x a0 -0.179 0.994 0.0 0.0 0.0 o-0 0.411 -0206 -0570 -0_043 O-057 OJXS o-0 O-0 o-0 O-071
6532 O-697 -0-614 0.0 01) 0391 0.0 0.0 0.0 01) QICt4 03 02&l O-053 -0_056 -0-327 0.179 0.0 o-0 o-0
vibrations of cytosine in this study, only the Raman and infrared bands assignable to the in-plane vibrations are given in table 4 and in fig 2 We can distin,puishbetween in-plane and out-of-pIane vibrations based on our present caktdation Our distinction is mostIy the same as that propo&d by Susi et al (12_14&Thus, markedIyb-mad and strong infrared bands at 823 and 520 cm-’ of cytosine and those at 605 and 388 cm-’ of cytosinel.amino+ are reasonably assigned respectively to NtH out-of-piane bending, NH2 wagging NrD out-of-plane bending, and ND_ wagging viirations With referenceto an anisotropy study of the infrared absorptions in a single crystaI of l-methyltbymine [15].568 and 442 cm-’ bands of qtosine and 550 and 438 cm-t bands of cytosinel,aminti3 are -ascribed to out-of-plane modes_ These are alI removed in table 4 and in fig 2 In additioa,twowetakRamanbands atlOlOand994 _-’ of cytosine and two Raman bands at 1046 and 1005 cm-t of cytosine-l,amincAs conId be assigned to CH out-of-plane viirations_ Therefore, th~WeredSOremove&
O-743 -0.097 -0.126 o-0 0.0 0.0 01) 01) 0.0 -0342 0.0 O-0 0_055 O-044 -O_lS-J O_I20 0.0 0.0
s_4i3 o-0 o-0 -0-149 o-0 0.0 --a103 o-0 -0_192 0.106 -0337 -O_l% -0.137 -0.065 0.0 -0113 0.0 0.0
5.169 0.0 0.0 O-0 o-0 o-0 o-0 o-0 0.0 o-0 o-0 O-0 0.0 0.0 0.0 o-0 0.0
13347 o-0 0.0 0.0 o-0 0.0 0175 0.199 -0308 0.0 0.077 0.0 0.0 0.0 o-0 0.0
8.452 o-0 o-0 0.0 0.0 0.186 -0302 0.0 o-0 -0_06s 0.072 0.0 o-0 -0_170 0.061
S-453 -a192 0.0 0.0 0.0 0.0 0.0 01) o-0 0.079 0.0 o-0 0.055 O_OS6
By an extensive test of the ab initio force constants [l-3,5-6,8.16-21). it is now generahy claimed that the 4-31G force constants of a moIecule can predict a correct in-plane vibrational mode spectrum_ When the calculated modes of a molecule are arranged in the decmasing order of the calcuhxted frequencies, along the arrangement of its observed fundamental bands in the decreasing order of the observed frequencies then one can reliably comxct one-by-one the members of these two arrangementswithout any cross connection- It is true that each of the caLdated normal frequencies comes out to be lo-20% higher than the corresponding observed fundamental frequency. Thisis partlybecauseoft!~eanhaxmonicityofthe vibration and partly &cause of the inability in the present MO calculation to incIude proper configuration interaction [20]. Nevertheless. the viirationaI modes. isotope effects, and infrared intensities of a moIecuIe are considered to be predicted by the 4-31G HF SCF MO method with a significant pre&iotL Based upon such a -generaI reliability of the
8.435 0.0 0.0 0.0 o-0 o-0 O-0 0.0 -0.059 0.0 o-0 0.03s -0.072
6.337 0.0 O-0 o-0 o-0 0.0 o-0 0.0 o-0 o-0 o-0 0.0
6.173 0.0 0.0 0.0 O-0 o-0 0.0 0.0 0.0 o-0 0.0
l-843 o-0 o-0 0.0 0.035 0.0 0.0 0.0 o-0 o-0
1.729 o-0 o-0
0.173 -0.045 0_087 -0_096 o-0 0.0
1.740 0.132 0.0 -0.119 0.059 o-0 o-0 0.0
o&O1 -0.060 0.0 O-014 o-0 o-0 0.0
1336 -0.041 0.0 -0.027 o-0 0.0
1.352 0.0 -0.020 0.036 0.193
0590 0.0 0.0 0.0
0.635 a.0 0.0
0574 -0.014
0.693
4-31G
calculation, the observed Raman and &Ifrared bands are assigned as shown in fig_ 2_ There
stretching vibrations, respectively_ In the 1800-850 cm-’ regiow the highest two
are a number of aiterations necessaq from the assignments proposed by Susi et al. 1121. Two strong infrared bands of cytosine observed at 3380 tid 3169 cm-’ are assignable to the NHI antisymmetric and symmetric stretching vibrations, respectively, and those of cytosine-dz at 2545 and 2337 cm-’ to the ND2 antisymmetric
observed frequencies, 1694 and 1653 cm-‘, are to be assigned respectively to the Cz=O and G% (or v”,) stretching vibrations_ Our cakulation indicates that the next highest frequency should be that of the NH, scissoring motion, and the strong infrared band at 1615 cm-’ corresponds to it_ It is
and symmetric stretching vibrations, respectively_ A fairly strong Raman line at 3117 cm-’ and a
moved down to 1182 cm-’ on deuteration- The next three highest frequencies should be assigned to v,” of the hexagon, C,-(NH,)
stretching, and
changed on NI and amino deuteration and are assigned to two CH stretching vibrations. Our calculation indicates that the N,-H stretching vibration should be Ioca~&_ at + slightly higher frequency than that of the NH, Metric stretch-
N,H in-plane bending vibrations in turn; and these are correlated respectivelywith the obserred 1533, 1498, and 1462 cm-’ Raman lines. On deuteration, the fit two remain nearly unchan~~ &iIe the last one (N,H b&d_) is moved to 950 ~cm-.‘_ The cakulation instructs-&at the GH.- GH in;
ing vibration in an isolated molecule_ It k.know& [22]. however, that a hydrogen-bonded irpino group gives often a broad kand at a much lower frequency. The broad infrared band of ‘cytosine around 2700 cm-’ and that of cytosine-d, around
ph&e bending vibration should come next, followed by the KekulC vibration, and next the GH GH lSO”-out-of-phase bending Correspond+g to these, w prominent Ramv Iinks are observed &t 1361, 1276, &id 1247 cm-‘, req+ctiv~ly._ T+
weaker one around
2150 cm-’ -
3050 cm-’
both remain un-
can be assigned ;o the N,H and wID NH, -. ~.I. ~. . .:-_ ~.~. .: .I
ro&ng vibration is predic+i to be the r&t _:_
Coicd.
Il. id 1
L
-_
-a
l-__ -_
l__
__
__
-_
s_ -_
y
.E
Obsi
E
c
R,R
--_
--___ --__ --_-_
_s-
__-
-
-_
:
S.
.
--.
. :
;-,
II
.I
-
--
:=__
:
:
II
a
I
I’i ‘I .
I
I
.
s . . . --. .-_
:-t__
-_z_=_ :
-_
Ill
I .
I
I L
.
:
iI
I
I II
I
9
T
2z
2
1864
1782
Cl633)
Cl6l2)
1645
Cl4981
I575
(14621
1083
(971)
_
639
(5971
.. . .
381
t4001
._
--.
1699
344'(369)
(1514)
highest
one, and the ll&
.&I-~
Rkxan Iin
assignabIetoit_.The threelowest-frequencyyibqtticnsin the 1800-850 cm-' range ai 1108,_971, T@
@
WI2 tot0
and 894 cm-’ are assignable to three-r&g vii& : $ons, v&_ Tr, ana v&, i&pec+Iy_ 1-nr+se three ,~ vibrations, pronounced couphngs unth the -NH, r&g mqtion are found (see -thecalculated 1197,
;:,1485
--
-
ring vibrations, On deuteratiqn, the an+io rocking vibration comes d&k to 850 cm-’ (fairly-s,trong infrared band),- and. a& is shifted up to 1021. a+-*_ In ad&ioq. v& is pushed up. 3~ hi&; as .. 1383 _cm-:_ throkgh +_mu@ing with the N,D hepd; ., .~ing niotioi -Itie ~K@ctxl* motion, 0; ,+:~othei_~ up 06 deute,mti&, fro@. ..
‘.
.-
...
-, :.
:
._.. . . .-
_ ~.._ ...._: _.. --
3 4 5 6 7 8 9
4013 3sM 3803 341s 3375 193il IS64 US2 1722
IO 11 12 13 14 15 16 17 18 19 20 21 22 23
Ia5 I575 1476 1364 1309 1241 1197 19S-s 988 S2L 639 572 567 3SL
1 2
NH,
PSU-
3354
N,H* NH, s str GHSW GH= c=o sv q=c,str NH2 sci d &N Str 8N,H 6CH+ K&d& 6CHNH2 r G”, -r&-i da bmtb $ sbm SCN
_ _
3176 3117 3050 1694 3653 1612 1533 14% 1462 1361 1276 1247 1x48 1108 971 894 792 597 546 533 400
3350 2700 3169
1703 1662 16i5 1538 1505 1465 1364 it77 1236 1150 1100 966 793 600 549 533
3418 3375 2978 2833 2751 1923
2388 1648 15% 1514 1491 1383 1316 1282 11% 1130 1021 965 --
1855 1699 1653 1521 1444 1366 12SO 1259 1123 1@aS1aM IsG9 Sll
2545 2150 2337 1637 1605 1517 1505 1377 1316 12Sl 1182 1120
849 m 590 524 504 369
628 555 541 344
=’ E$Ihcuscofrtief--~tsgi~znintile3_ ” Rcf_ [9f
tiv&y_ All of these Four are moved narrowly towards lower frequency on N, and amino deuteration_
4, IxscusGon In our present study- the C,-amino stretching Frequency (1505 cm-‘) of cytosine is Found to be of interest_ This is extraordimniIy high in comparison, For exampI< with the C-amino stretchingfrequency (1276 cm-‘) of anihne [23]. This C-N bond is considered to have the greatest doublebond character among the C-(NH,) bonds of nucleic acid bases (cytosine, adenine, and guanine) [24]_ The height OF the barrier in the >C-NH, internal rotation decmases in going from cytosine, to adenine, and guanineAnother point OF interest in the cytosine mole-
cule
is its
“Kekdi
structure?_
As is seerk in the last
of table 1, the six bonds Forming the ring are long and short alternately. The bond-stretching Force constant changes also up and down ahernately on going along the six-membered ring (see the first six diagonal elements shown in tabIe 3) Thus, the bond order is considered to become higher and lower on going along the ring ThereFore, the KekuIi viiration (Kk), in which every bond in the six-membered ring stretches and contracts with a 180° phase difference from the stretching and contraction of the adjacent bond, may cause a gre& over&I poIari%abiIity oscihation in cytosine, whereas it should never in benzene_ Actually, the Raman line at 1276 cm-’ assigned to Kk of cytosine is very strong (fig 3) with the 488.0 nm excitation. The assignment of Kk in the cytidine residue is not made as straightForwardly as for cytosine itcohmn
.
-
:
-.
self_ -In-the Raman -.spectrum~of 5’CM.p
or 5’-
dCMP, there are two Iin&-at 1290 and.1250 cm-‘, and.& a. broad band %&nd 1230 Cm-‘. The band a&md 1230 cm-1 is .as&nabie -to 6CH-. The fint two Raman lines are considered to be originated by the &In-ationaI coupling between the Kk .motion. of the cytosine moiety -and a sugar vibration. Considering~ the intensities of the&e two Raman Ii&s in the visible excitation, however, we prefer to assign the 1250 cm-’ Line to a mode in which Kekule motion is predominant. The KekuIe structure of cytosine in its ground electronic state must .be-disorganiz& -in its first electronic excited state at 268 nm [25]_ As is seen in fig_ 6, on going from the highest occupied to the lowest vacant molecular orbit& the Ns=C, and Cs=C, bonds should change from bonding. to antibonding; while the C,-N, and C,-C, bonds should change from antibonding to bondinK, the bond order decreases in the former two and increases in the latter two, and therefore G-N, must be shortened, N,=C~ lengthened, C,-C, shortened, and C,=C, lengthened_ Thus, the geometry change in the lo&st electronic transition may rather match the Kekule motion_ Therefore, the Kekult vibration around 1250 cm-’ of the cytosine residue must be active in its resonance Raman spectrum with excitation in its 268 nm absorption band. This is found to be actuahy the case. The 1243 cm-’ Iine of S-&BP and the 1255 cm-’ Iine of S-dCMP are very strong in their Raman spectra excited at 257.3 run [26,27] or at 266 nm [28-301. It is noteworthy, on the other. hand, that they are completely absent with excitation at 240, 218, or 200 nm [28-301. The. geometry of the cytosine residue in the second- and the third-lowest excited states does not. seem. to match the KekuIi type deformation, The NH, and ND, scissoring viirations have been Iocated in our present study at -1612 (cytosine) arid 1190 (cytosine-cZs) cm-‘, respectiveIy_ This supports the assignment of the 1607: cm-’ Raman Iine of 5’dCMP (in aqueoussolution) to its NH, scissoring .vibmtion made recentIy by Fodor et al [30]. To its ND, scissoring, we assign a Raman Iine at 1181 -cm-?-of S-dCMI? in l&O sohrtion~ The 1607 ‘-cm-l line. ‘of 5IdCMP w& found to be enhanced by ihe 200 inn excitation
transition from a u orbital to the 3s orbit$;of&e' nitrogen atom around 2O(I run. Such a’transitiori_.. should have its transition moment _&mg the-dir@. tion (2) perpendicular to the rnol&rl&~pIa&<~~@id me the zz component of the Raman sc%tteririgte&& 1 of the NH, .scissoring vibration mi&t-be m-r&~:-
n&e -5th &is &&o~c t.ra,n.sition_.-. _-Z-~-.ci ._:- _I’ -As discuss~ above, the present viirationd assignmentsfor the cytosine molecule &e often ;Seful in the vibrational &gnments for & cytidine derivative_ S-dCMP, for example, shows -weaker Raman scattering in the low-frequency &de of the strong 1255 cm-’ Iine (Kk) ~withpeaks at 1242 ’ and 1214 cm-’ 1301.The 1242 cm-’ is assignable to SCH-, but the 1214 cm-’ line‘& not attributabie to any_ of the-vibrations of the-base moiety. ] Hence this is suggested to be caused by. the ‘de_ oxyribose moiety_ As mentioned in connection with the assignment of the Kekule vibration above, we may expect a few vibrations of the sugar in the 1200-1300 cm-’ range_ Indeed, we have recently found the spectraI feature in the 1300-1200 cm-’ range of cytidine is sensitive to the. deoxyribose conformation_ Some vibrational couplings between them would cause such a conformation dependent
Fig 6. ‘Ihe highest occupied and _lowes< va&ant ~m&klar orbita& of the P el&ns in thecytosine mokcul~ on the basis of & ~publi+hed part of the result&of a P&iser+axT-Pople calculation made by Nagata et al [25]_ Open w$ closed .tie~ idicace that the upper bra&he+ of the ato’&& orbit+ of the.= ekctrons iic &sitive And nk@iv~$n+ectively_ l-li~_s+e of the &de indicates thk appr&+ate value of the.coetXcia;t of th+ par&lar atomic orbital in .&e relevant moIe&+~ital. a&zre (rad;u~)~ E coefkitnt .~
~_ ~.:
-.
spew-al feature- The details w-ill be given in our comingpaper-
Our. sinaxe thanks are due to Professor K. Morokuma, Institute for MoIecuIar Science, and Dr_ S_ Kate, Nagoya Utiersity, for their kind arrangements and instr~.~ctio~~~ in our ab initio MO cahhion with the ILMSPAC progg Tf?e normaI frequency and normal mode caiadation was carried out at the Computer Center of IMS (HIIAC M-ZGOH)_ The pIotter output of the normal modes was made at the Computer Center, University of Tokyo (HITAC M-20OH)_ This work u-as supported by a grant (No_ 57060004) from the ~finistry of Education. !Zciace. and Culture of
Japan References Y_ Pikhimm
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CW_ Bock, M. Tmcband P_ Gcorgc J. MoL Spatry_ SO (1980> 131; 84 (19sO) 243.2s [191 f_ PkIay and W_ Meyer. J_ hid Specu-y_ 40 (197l) 59; W_ Meyer and P_ P&y. Theoret Chin Acta 32 (1974) 353: P. Ptday and F. -I’&&. J. Mol. Struct 29 (1975) 239; P_ P&y_ G_ Fogamsi. F_ Pang and J-E- Boggs J_ Am_ Cban Sot 101 (1979) 2550: P_ Pulay. G_ Fogam& G. Pongcr. JE Boggr and h Vzqa, J_ Am Cbcme Sot 105 (1983) 7037_ [201 N_ Tanz&a. Y_ I-Izmzda. Y_ Suga~swa. kM_Tsuboi S Kato and K_ hfot-okxmsa. J. MoL Spatry_ 99 0983) 245; N_Tanaka,Y_H-daandM_Tsuboi,Ckm_Pbyr94 (1985) 65. [IIf Y_ Sugaaarq A_Y_ Hirakawa and M_ Tsuboi, J_ MoL spa=uy_ 10s (1984) x6_ 1221 CL An@. I Chau Sot (l%l) 5w. 1231 M_ Tsuboi. Spafmchk Acta 16 0960) 505_ [24] M_ Nakau&i. S. Tstmcda. S. TakahasE and hi. T_suboi.J. Biodrem (Tokyo) 97 (3985). to be published_ (251 C Nagam. A lmamura and H Fujira in: A&anccs in biophysics. VoL 4. aL M Kotani (Uniuazity of Tokyo Press Tokyo. ‘-mj pm1. [26] Y_ Nkbimma. AYe Hirakana and U Tsuboi. C&em_ LcIKux(l977.)907_ [2q Y_ Nishimm-& A_Y_ m and M Tsuboi, in: AdtanccsiItinfrarrdand~~roscopy*voL5,cds_ RJ.Ha Clark and RE Hcncr (Heyday, London, 1978) p_ 212 1281I-D. Zicglcr. -B Hudson. D-P. Screw and W_I._ Pc?icok!s,Biopolymas 23 (1984) 2067_ [zsl WX Kubasdc. B_ Hudson and WE PcticoIas, Pmc NatL ~Acxd.Sci_US.cobekpublisbed. [30] S_P+ Fodor, RP. Rava aiid T-G: $& J_ AI+ C+_ sot 107 (1985) 1520_